Platelet transfusions are increasingly being administered to prevent bleeding in many clinical situations. Unfortunately during storage platelets lose viability such that their hemostatic effect becomes diminished. The added risk to patients and expense is difficult to calculate but some have estimated that as many as one-third of the 6 million platelet products transfused each year are of marginal quality. The biochemical and molecular events responsible for this "storage defect" have been poorly characterized. This proposal takes a novel approach to understanding the loss of platelet viability during storage by applying the concept of programmed cell death (PCD) or apoptosis to platelets. Programmed cell death is an intrinsic mechanism in all cells triggered by many stimuli. The hallmark events include loss of Bcl-2 and mitochondrial function, release of cytochrome c and consequently activation of the caspase system, exposure of phosphatidylserine (PS) on the cell surface, and degradation of the nucleus. Preliminary work described herein shows that the presence of Bcl-2 and several crucial members of the caspase family in platelets and that caspase-3 is activated in platelets during storage. This proposal seeks to define the PCD pathway in platelets and then find inhibitors of PCD that might ameliorate the platelet storage defect. Four goals are proposed: First, the PCD pathway in platelets will be fully defined as for the phosphatidylserine surface membrane exposure and platelet-specific synthesis of apoptotic proteins. Second, the changes in these members of the PCD pathway will be characterized in detain over 5 to 7 days of storage. Third, studies will be performed to determine whether elements of the platelet storage environment can trigger PCD. Finally, since apoptosis can be inhibited by growth factors (such as thrombopoietin and stem cell factor) or small molecules (such as the caspase inhibitors DEVD-CHO and Z-VAD.FMK) even in the presence of ischemia, these will be tested for their ability to prevent PCD in platelets during storage. Such interventions have successfully improve the viability of other cell types and may do the same for platelets. Successful conclusion of these studies will lead to a clearer understanding of the molecular regulation of platelet viability and prove a means to improve it.